A multidisciplinary approach will be used to determine the mechanism of lysosomal movement and enzyme mobilization in human polymorphonuclear leukocytes (PMN). This approach involves the use not only of the lysosomal system in PMN themselves, but other closely related particle translocation and release systems, including pigment granule (melanosome) movement in melanocytes, and release of neurotransmitters and associated enzymes from synaptic vesicles. The motive force behind the translocation of lysosomes, in relation either to phagosomes or to the cell surface, will be studied by ultrastructural and biochemical analysis of lysosome-associated filments, both in resting PMN and in PMN stimulated by the plant lectin concanavalin A or by opsonized particles presented for ingestion. The work will include the identification of 7nm filaments as actin, both in situ in cells made permeable to high molecular-weight reagents, and in vitro on isolated lysosomes. Further studies will include subfractions of human lysosomes in which the coisolating proteins--and especially the putative contractile proteins--will be examined. We will also study the importance of microtubule integrity on the synthesis of cyclic AMP and cyclic GMP, second messengers thought to regulate lysosomal enzyme mobilization in PMN> The possible role of protein phosphorylation in this regulation will be studied by examination of the phosphorylation of endogenous proteins in both intact and broken cell preparations. In addition, as a model for the relation of protein phosphorylation to enzyme release, we will examine the effects of colchicine, cytochalasin B and related drugs on neurotransmitter release, cyclic AMP levels, and protein phosphorylation in synaptosomes. Finally, particle translocation will be studied in frog dermal melanocytes, in ATP-reactivated particle movement systems similar to those being used to study sperm flagellar motility and anaphase movements of chromosmes in the mitotic apparatus. This type of systems should permit a direct demostration of the contractile apparatus involved in particle movement, and would be a unique, functional, reactivated model for considering particle translocation in general.